Competitive inhibition by substrates of the esterification reaction between l-phenylalanine and d-glucose catalysed by the lipases of Rhizomucor miehei and Candida rugosa

• Published in: World journal of microbiology & biotechnology 2007-07, Vol.23 (7), p.955-964
• Main Authors: Lohith, Kenchaiah, Manohar, Balaraman, Divakar, Soundar
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biotechnology; Candida rugosa; Candida rugosa lipase; Competition; Competitive substrate inhibition; Computer simulation; Curve fitting; d-Glucose; Dead-end lipase-substrate complex; Esterification; Fundamental and applied biological sciences. Psychology; Glucose; Kinetics; l-Phenylalanine; Phenylalanine; Ping-Pong Bi-Bi mechanism; Proteins; Rhizomucor miehei; Rhizomucor miehei lipase; Substrate inhibition
• ISSN: 0959-3993; 1573-0972
• DOI: 10.1007/s11274-006-9320-4

A kinetic study on esterification between d-glucose and l-phenylalanine catalysed by lipases from Rhizomucor miehei (RML) and Candida rugosa (CRL) in organic media investigated in detail showed that both the lipases followed a Ping-Pong Bi-Bi mechanism with two distinct types of competitive inhibitions. Graphical double reciprocal plots and computer simulation studies showed that competitive double substrate inhibition took place at higher concentrations leading to dead-end inhibition in the case of RML and in the case of CRL, inhibition only by d-glucose at higher concentrations leading to dead-end lipase-d-glucose complexes. An attempt to obtain the best fit of these kinetic models through curve-fitting yielded in good approximation, the apparent values of important kinetic parameters, RML: k cat = 2.24 ± 0.23 mM h-¹ (mg protein)-¹, K m l₋phenylalanine = 95.6 ± 9.7 mM, K m d₋glucose = 80.0 ± 8.5 mM, K i l₋phenylalanine = 90.0 ± 9.2 mM, K i d₋glucose = 13.6 ± 1.42 mM; CRL: k cat = 0.51 ± 0.06 mM h-¹ (mg protein)-¹, K m l₋phenylalanine = 10.0 ± 0.98 mM, K m d₋glucose = 6.0 ± 0.64 mM, K i d₋glucose = 8.5 ± 0.81 mM.